tbuckley/cabot-search huggingface dataset:
Abstract P3165: Spectrin-based Regulation Of Exosome Secretion In Cardiac Fibroblasts
Authors: Rebecca J Shaheen, Evelyn Thomas, Drew Nassal, Nehal J Patel, Xianyao xu, Thomas J Hund
Journal: Circulation Research (2023)

Cardiac fibroblasts (CFs) not only modulate the extracellular matrix under physiological conditions, but also have the ability to sense local injury and respond with a phenotypic change and increased paracrine communication to distal cells. While a myriad of studies have focused on identifying the paracrine signals involved in recruitment and activation of CFs, the manner by which CF paracrine communication networks are tuned in response to chronic stress remains unclear. Previously, our lab found that stress-induced loss of the cytoskeletal protein, β IV -spectrin, regulates the CF phenotype and fibrosis. Therefore, we hypothesized that degradation of β IV -spectrin is a critical step in stress-induced communication involved in recruitment of distal CFs. In this study, we first collected conditioned media from WT and β IV -spectrin deficient (qv 4J ) CFs to test the effects on quiescent WT CFs. We found that the CFs treated with qv 4J -conditioned media showed increased proliferation and compaction rates, as well as altered gene expression in the recipient cells compared to those treated with WT-conditioned media. To further explore the role of β IV -spectrin in paracrine signaling, we isolated exosomes from WT- and qv 4J -conditioned media. We found that loss of β IV -spectrin in CFs resulted in an increased concentration of exosomes in conditioned media, that also induced increased proliferation in quiescent CFs. Furthermore, we found that genetic mice expressing degradation-resistant β IV -spectrin ( qv 3J mice) showed impaired CF recruitment and activation following myocardial infarction (MI) together with increased mortality and impaired fibrotic remodeling compared to WT. Finally, we demonstrate that exosomes from a WT-MI heart can improve fibrotic remodeling in qv 3J mice following MI. Our work identifies a novel role of the spectrin-based pathway in facilitating long-range communication in response to chronic stress.
Abstract GS.07: Cardiac Bridging Integrator 1 Gene Therapy Rescues Heart Failure In Swine.
Authors: Jing Li, Pia Balmaceda, Thuy Ha, Eugene Kwan, Tara Hitzeman, Joseph Palatinus, Guillaume Hoareau, Ravi Ranjan, Robin M Shaw, Tingting Hong
Journal: Circulation Research (2023)

Heart failure (HF) is a major cause of morbidity and mortality worldwide, yet with limited therapeutic options. Recent evidence indicates that the cardiac bridging integrator 1 (cBIN1)-organized transverse tubule microdomain is key to calcium handling and cardiac function. In rodent models, adeno-associated virus 9 (AAV9)-cBIN1 effectively reconstitutes microdomains to rescue HF. Here we explored the therapeutic efficacy of AAV9-cBIN1 in a minipig model of non-ischemic HF. 6-7 months old Yucatan minipigs were subjected to continuous right ventricular rapid pacing (RVP) at 170 bpm. After 8-11 weeks of RVP, minipigs developed dilated cardiomyopathy with increased left ventricular (LV) end diastolic volume (EDV, 69.0 ± 5.3 vs. baseline 50.0 ± 3.8 mL) and reduced ejection fraction (EF, 36.3 ± 2.9 vs. baseline 68.7 ± 0.8 %). Once EF ≤ 40% over a week, pigs were randomized to receive AAV9-cBIN1 (6 x 10 11 vg/kg, intravenous, n = 3) or control (AAV9-GFP or PBS, n = 3). Weekly echocardiography, bloodwork, clinical assessment, and pacing interrogation were performed. In control animals, LV further dilated with EF declining and all three animals died of severe HF symptoms by 7 weeks post injection (terminal EF 18.3 ± 1.7, EDV 107.4 ± 25.0). Despite of continuous RVP, AAV9-cBIN1 limited LV dilation and rescued EF (58.0 ± 4.2, 59.0 ± 4.4 % at 4, 13 weeks post injection). Two cBIN1-treated animals survived to the end of protocol at 6 months post treatment (EF, 57.5 ± 15.5 %), with the third pig currently surviving at 13 weeks post treatment with a 60% EF. No significant toxicity was observed. In conclusion, low dose intravenous cBIN1 gene therapy rescues non-ischemic HF and improves mortality in pigs.
Abstract P3180: Cardiomyocyte Pgc-1α Mediates The Adaptive Response To Endurance Exercise Training In Mice
Authors: Sumeet A Khetarpal, Haobo LI, Claire Castro, Nicholas H Houstis, James Rhee, Hans-Georg Sprenger, Melanie Mittenbuhler, Alexandra Kuznetsov, Rebecca Freeman, Phillip Dumesic, Jason D Roh, Anthony Rosenzweig, Bruce Spiegelman
Journal: Circulation Research (2023)

Exercise has myriad systemic benefits in reducing the incidence from and morbidity of cardio-metabolic diseases. In healthy individuals, aerobic exercise promotes physiologic hypertrophy of the heart, which requires increased mitochondrial numbers and function. In skeletal muscle and heart, a key orchestrator of the adaptation to increased energetic needs is the transcriptional regulator PGC-1α. PGC-1α may mediate cardiac adaptation to physiological stress through its function in mitochondrial biogenesis but also through other mechanisms. The role of cardiac PGC-1α to the systemic training response is poorly understood. We generated PGC-1α cardiomyocyte-specific knockout (cKO) mice to study its role in endurance exercise training. Sedentary cKO mice demonstrated normal cardiac function and no significant differences in acute treadmill exercise tolerance. However, cKO mice demonstrated a lack of ability to augment exercise capacity after 6 weeks of wheel running compared to WT (work achieved 32.9 J in WT [39% increase] vs 23.1 J in cKOs [no change vs baseline], p&lt;0.001, unpaired t-test). This was despite comparable voluntary wheel distance run during training (2.93-3.67 km/day for both over 5 weeks, p=0.58). Unexpectedly, after 6 weeks of training, cKO mice developed a dilated cardiomyopathy at rest and exercise-related dysfunction (contractile reserve [ΔFS stress-rest ] +6.0 in WT vs -6.4% in KO, p&lt;0.05, unpaired t-test). They also demonstrated features of cardiac cachexia as measured by 30% and 21% reductions in inguinal adipose and gastrocnemius mass, respectively (P&lt;0.05, unpaired t-test). Gene expression profiling demonstrated activation of pathological ( Nppa , Nppb , Myh7/Myh6 ) and atrophic pathways ( Gdf11 , Gdf15 , Fstl3 ) in the trained cKOs. In vitro, gain of function studies using adenoviral expression in neonatal rat ventricular myocytes demonstrated that PGC-1α overexpression could abrogate pathologic hypertrophy and atrophy gene expression pathways stimulated by phenylephrine. Our preliminary studies support cardiomyocyte PGC-1α as a key mediator of the adaptive response to exercise training and unveil a key role of this transcriptional activator in protecting from exercise-related cardiac dysfunction in mice.
Abstract P2066: Forced Exercise Prevents Reductive Stress Induced Cardiomyocyte Remodeling And Cardiac Dysfunction
Authors: Arun Jyothidasan, Sini Sunny, Silvio Litovsky, Steven M Pogwizd, Rajasekaran N Namakkal-Soorappan
Journal: Circulation Research (2023)

Background: Disruption of the endoplasmic reticulum (ER) function has been recognizedas a clinical signature of congestive heart failure, ischemia, and atherosclerosis. Whilepro-oxidative redox milieu is vital for the ER to regulate protein folding events. Recently,we reported that overabundant antioxidants in the heart cause reductive stress (RS) andimpair the protein folding mechanisms due to unresolved ER stress. Here, we testedwhether forced exercise prevents RS and improves ER function through augmenting ROSsignaling, thereby improving the PQC mechanisms in the heart. Methods: Cardiac-specific constitutively active Nrf2 (CaNrf2-TG) and non-transgenic (NTG) mice at 5-6weeks of age were subjected to forced treadmill exercise (FTE) for 20 weeks (15meters/minute with a 5° slope; for 45 minutes/day; for 6 days every week). Cardiacstructure and function were analyzed before and after the exercise. At 22 weeks, micewere sacrificed for analysis to assess the benefits of FTE. Results: RNA sequencingrevealed significant downregulation of key protein folding genes and chaperones in TGmice in association with the pathological remodeling at 6 months. Particularly, Pdi, Atf6, Calr, and Ero1α were downregulated (&gt;2 fold) at 24 weeks in TG mice, indicating impairedER function in the remodeling hearts. These changes were coupled with increasedcardiomyocyte size and hypertrophy. Upon FTE, we observed significant improvementsin the structure and function of the cardiomyocytes. WGA staining showed the cross-section area of the cardiomyocytes was significantly reduced in the exercise groupcompared to the sedentary-TG mice experiencing cellular hypertrophy (115.7μm 2 NTGvs 428.8μm 2 TG vs 202.5 μm 2 TG-FTE). Echocardiography analysis demonstratedpreservation of cardiac structure and function (EF, FS, IVD, IVS, EDV, and ESV).Specifically, compensatory improvement in diastolic function (MVE/A ratio) was observedin TG-FTE exe group. Additionally, we observed that the ubiquitination of proteins wassignificantly reduced in the exercise group indicating deaccumulation of the misfoldedproteins in RS hearts. Conclusion: Together these results demonstrate that chronicexercise could ameliorate RS-induced ER stress and cardiac dysfunction.
Abstract P1121: Substrate Rigidity And Mybpc3 Genotype Regulate Development Of Hypertrophic Cardiomyopathy Structural And Functional Phenotypes In Human Ipsc-derived Micro-heart Muscle Arrays
Authors: Jingxuan Guo, Huanzhu Jiang, David Schuftan, Lavanya Aryan, Nathaniel Huebsch
Journal: Circulation Research (2023)

Introduction: We hypothesized that mechanical loading is a key factor in the early stages of hypertrophic remodeling associated with pathogenesis of hypertrophic cardiomyopathy (HCM). We tested this hypothesis in vitro with human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CM) bearing heterozygous loss-of-function mutations in Myosin Binding Protein C (MYBPC3). Cardiomyocytes were cultured in engineered micro-heart muscle arrays that were loaded by changing substrate rigidity. Methods: Isogenic control and MYBPC3 +/- hiPSC-CM were seeded into dogbone-shaped molds to form micro-heart muscle atop silicone rubber substrates with rigidity levels ranging from levels of embryonic (0.4 kPa) to fibrotic adult heart (114 kPa). Spontaneously contracting micro-tissues formed within 48 hours. 10 days after formation, tissues were monitored for action potentials (voltage sensitive dye, BeRST-1), Ca 2+ transients (GCaMP6f reporter), and contractility (substrate deflection). Cell and tissue architecture were assessed in 8 μm cryosections, and RNA transcription via qRT-PCR. Results: MYBPC3 +/- and control cardiomyocytes had similar physiology and morphology in 2D monolayers. However, MYBPC3 +/- tissues exhibited marked changes in Ca 2+ handling, which were exacerbated when substrate stiffness increased to the level of healthy heart muscle (15 kPa). This was concurrent with hypertrophy of MYBPC3 +/- cardiomyocytes at 15 kPa. MYBPC3 +/- tissue Ca 2+ handling abnormalities were linked to enhanced Ca 2+ intake rather than SERCA deficiency or excessive myofilament buffering. MYBPC3 +/- tissues exhibited mis-regulation of cardiac troponins on the RNA and protein localization levels, which may underlie the observed structural and physiological changes. Conclusions: Changes in the rigidity of the substrate, mimicking changes in heart muscle caused by blood pressure, pushed hiPSC-cardiac micro-tissues with MYBPC3 mutations to recapitulate key structural and electrophysiological characteristics of HCM. In contrast, control tissues exhibited hypertrophy and abnormal calcium handling, but only at extreme values of substrate stiffness. This suggests that HCM mutations may exaggerate load-dependent hypertrophic remodeling.
Abstract P2075: The Effects Of Maternal Psychosocial Stress During Pregnancy On Offspring Heart Health
Authors: Sarah L Sturgill, Mark T Ziolo, Lackritz E Taryn
Journal: Circulation Research (2023)

Maternal health has a substantial effect on offspring well-being and health. A significant contributing factor to maternal health is psychosocial stress. Psychosocial stress results from adverse experiences in everyday life and the inability to cope with them, resulting in anxiety and depression. Unfortunately, it is estimated that ~75% of mothers will suffer a stressful life event the year their infants are born, and upwards of 20% suffer depression. These already high levels of psychosocial stress have been exacerbated by the COVID-19 pandemic. However, it is not known if maternal psychosocial stress during pregnancy affects offspring heart health. Thus, the purpose of this study was to investigate the effects of maternal psychosocial stress during pregnancy on the heart health of offspring. We hypothesize that maternal psychosocial stress during pregnancy will have an adverse effect on offspring heart health. Methods: We used a well-established model of maternal psychosocial stress during pregnancy- restraint stress. Pregnant dams were stressed by placing them in a 50 mL conical tube (with air holes) for 2 hours / day on gestational days 10-17. Offspring heart health was assessed via echocardiography (Vevo 3100, MX550D transducer). Results: As early as one week of age (equivalent age of infancy in humans), we observed that offspring born to stressed dams exhibited decreased left ventricular fractional shortening. Further analysis using speckle tracking echocardiography was performed on offspring at 4 (equivalent age of juveniles in humans) and 12 (equivalent age of young adult in humans) weeks of age. Our data indicate that offspring from stressed dams had decreased heart performance (measured as stroke volume), remodeling observed as hypertrophy (measured as left ventricular posterior wall thickness at diastole), and diastolic dysfunction (measured as E’/A’). Conclusion: Our data indicate maternal psychosocial stress during pregnancy results in blunted left ventricular contractile and diastolic dysfunction in offspring, as well as displays adverse remodeling (i.e., hypertrophy) during infancy and through young adulthood. These data indicate that maternal psychosocial stress during pregnancy is detrimental to offspring heart health.
Abstract P2112: Mechanism Of Bag3's Degradation Of Tgfbr2 In Cardiac Fibroblasts
Authors: Margaretha Morsink, Bryan Wang, Josephine Watkins, Lori Luo, Xiaokan Zhang, Rajesh Soni, Barry Fine, Gordana Vunjak-Novakovic
Journal: Circulation Research (2023)

BAG3 levels are diminished in patients with heart failure and several mutations have been identified to cause dilated cardiomyopathy with cardiac fibrosis. BAG3’s four binding domains are critical to its function to facilitate proteostasis. BAG3’s role has been extensively studied in CMs, where it aids sarcomere protein turnover through chaperone-assisted selective autophagy, through BAG3’s complex with HSP70, HSPB8, and E3 ligase CHIP. However, BAG3 is ubiquitously expressed and its role in non-myocytes has been overlooked. Cardiac fibroblasts are responsible for the excessive matrix deposition leading to fibrosis, mostly through TGF-beta signaling. Previously, we have shown the importance of BAG3 in cardiac fibroblasts through its role in TGF-beta signaling and identified BAG3 to facilitate the degradation of the type II TGF-beta receptor (TGFBR2). As BAG3 does not have inherent E3 activity, this study aimed to identify the specific E3 ligase of TGFBR2 and the binding domains of BAG3 that are crucial to this interaction to gain insight into the effects of the clinical mutations of BAG3. Immunoprecipitation (IP) of endogenous BAG3 in cardiac fibroblasts revealed five E3 ligases as high-confidence interactors of BAG3. To find the specific E3 ligase of TGFBR2, we performed an IP of a V5-tagged TGFBR2 and identified HSP70, HSPB8 and CHIP as binding partners. To delineate the critical domains of BAG3 that impact the TGFBR2 degradation, we overexpressed 3x-FLAG-tagged BAG3 proteins lacking each domain respectively (WW, IPV, PxxP, and BAG) and show that the BAG domain is imperative in TGFBR2 degradation. This has clinical implications for the common DCM-causing E455K mutation in the BAG domain. The ubiquitination of TGFBR2 was decreased upon overexpression of a BAG3 plasmid containing the E455K mutation. Lastly, the drug JG-98, which disrupts the binding of HSP70 to BAG3, was used to show the importance of the HSP70-BAG3 axis in the degradation of TGFBR2. In conclusion, we have uncovered the molecular mechanism of TGFBR2 degradation in cardiac fibroblasts mediated by the BAG3-HSP70-HSPB8 complex, with clinical implications for the loss of BAG3 and several common DCM-causing mutations in the development of cardiac fibrosis.
Abstract P1117: Personalized Care Using Patient-derived Cardiac Spheroids For Heart Failure Patients
Authors: Carmine Gentile
Journal: Circulation Research (2023)

Preclinical studies using currently available in vitro and in vivo models fail at fully recapitulating the in vivo human heart microenvironment and its pathophysiology, limiting their translation from the bench to the bedside. Our laboratory has developed cardiac spheroids (CSs) as advanced in vitro models of the human heart using cardiac myocytes, endothelial cells and fibroblasts at ratios approximating the ones found in the human heart. Thanks to patient-derived induced pluripotent stem cells (iPSCs), we have quickly moved to personalized CSs to establish advanced pathophysiological in vitro models of heart failure (HF). CSs have been used to evaluate short- and long-term effects of ischaemic/reperfusion (I/R) injury typical of myocardial infarction (MI), drug-induced toxicity, SARS-CoV-2 exposure and pregnancy-induced HF. This was achieved using our established protocols to induce I/R-mimic injury, as well by exposing CSs to doxorubicin (DOX), SARS-CoV-2 virus and pre-eclampsia blood samples, respectively. First, our analyses of cell toxicity ratios using calcein-AM and ethidium homodimer (staining live and dead cells, respectively) demonstrated treatment-specific loss in cell viability and increase in cell death compared to control (untreated, healthy) CSs. Other CSs stained with ethidium homodimer and cell-specific antibodies were imaged and further used for 3D rendering analyses using IMARIS software. This enabled us to identify cell-specific responses in each pathological condition. Then, our measurements of fractional shortening and contraction frequency in CSs showed a statistically significant loss of contractile function in treated CSs compared to controls. Finally, new potential targets to prevent myocardial injury were evaluated using transcriptomics analyses, with a focus on changes in expression levels for genes regulating sarcomere structure, calcium transport, cell cycle, cardiac remodelling and signal transduction. Current studies are evaluating novel potential therapeutic targets to prevent and treat myocardial injury in HF patients using CSs. This has the potential to pave the way to prevent and treat HF in a safer and personalised way, hopefully preventing undesired effects due to non-specific care.
Abstract P1100: The PD1/PD-L1 Pathway Provides An Immunosuppressive Environment Necessary For Neonatal Heart Regeneration
Authors: Stephanie Vargas Aguilar, Miao Cui, Ning Liu, Rhonda S Bassel-Duby, Eric N Olson
Journal: Circulation Research (2023)

The neonatal mouse heart regenerates after injury without scarring or impairment in heart function, but this ability is lost within the first week of life. This regenerative time window coincides with important changes in immune cell composition, maturation, and function. To understand how these changes support regeneration over scarring in the neonatal heart, we compared the immune response in regenerative and non-regenerative mouse hearts following myocardial infarction (MI). The immune response in regenerative hearts is characterized by a low degree of inflammation that is rapidly resolved, whereas non-regenerative hearts exhibit an exacerbated and prolonged immune response that involves strong T cell activation. Accordingly, we discovered that the PD1/PD-L1 pathway, an immune-modulatory pathway that acts mainly through the suppression of T cell activation, is highly activated in regenerative hearts but rapidly silenced during the postnatal period. Deletion of the PD1 receptor or inactivation of its main ligand, PD-L1, prevented regeneration of neonatal hearts after MI. The immune response in neonatal hearts from PD1 knockout mice was characterized by increased monocyte infiltration, a pro-inflammatory gene signature and exacerbated T cell activation. Our findings reveal a previously unrecognized role of the PD1/PD-L1 pathway in heart regeneration through modulation of the immune response and provide new insights into the importance and characteristics of a regeneration-permissive environment.
Abstract P2188: Covid-19 Plasma Exposure Causes Transcriptional, Morphological, And Functional Changes Within The Pulmonary Microvascular Endothelium
Authors: Rainha Passi, Justyna Cholewa-Waclaw, Ryan Wereski, Stefan Veizades, Matthew Bennett, Brownwyn Berkeley, Emmanouil G Solomonidis, Andrea Caporali, Ziwen Li, Abdelaziz Beqqali, Mieke Dewerchin, Nicholas L Mills, Mairi Brittan, Andrew H Baker
Journal: Circulation Research (2023)

Introduction and Aims: While primarily a pulmonary disease, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can affect multiple organ systems, including the vasculature. Thus far studies have successfully characterised the whole organ compartment, however, our understanding of the mechanisms associated with COVID-19 infection and its impact on pulmonary microvasculature (pMV) remains incomplete. In human pulmonary microvascular endothelial cells (HPMVEC) treated with plasma from patients hospitalised with COVID-19, we aimed to evaluate transcriptional, morphological and functional changes, and to provide a mechanism for the excess mortality in patients with COVID-19 and how it may inform therapeutic advancement. Methods and Results: Plasma acquired from hospitalised patients with COVID-19 was compared to plasma from patients hospitalised without COVID-19 but with other acute illnesses, such as COPD. Exposure to plasma from patients with COVID-19 caused a significant functional decline in HPMVEC, with a decrease in both cell viability on the WST-1 cell-proliferation assay (n=75, p= 0.013) and cell-to-cell barrier function measured by electric cell-substrate impedance sensing (n=3, p = 0.033) . High content imaging using the cell painting image-based assay showed a phenotypic change in nuclear (n=4, p=0.019) and nucleic acid (n=4, p=0.022) morphology, indicative of a deterioration in core biological processes including protein transcription and metabolic output. Comparative deep RNA-sequencing of HPMVECs treated with COVID-19 plasma (n=5) confirmed upregulation of genes, notably IFIT5 and SFMBT1 . These genes are known to negatively regulate transcription and respond to viral infection, respectively, further demonstrating the phenotypic and transcriptional decline in the pulmonary microvasculature in patients with COVID-19. Conclusion and Impact: Our findings suggest that core biological processes are disrupted in the lung microvascular endothelium during severe COVID-19 disease contributing to aggravated disease. From a translational perspective, this knowledge could provide insight into possible treatment opportunities and prognostic tools that may directly target the dysregulated endothelium.